The genetic variability and Individual variations in immune status dictate responses to vaccinations, infections, and contribute to disease severity. The sequencing of the human genome and generation of the Haplotype Map now enables a mechanistic understanding of how genetic variation influences human immune responses. Yet manifold non-genetic factors also interact to maintain the healthy immune system, and complex analysis will be required to form predictions for its response to perturbations. Here, we will employ systems approaches and novel, high throughput and high-fidelity technologies such as multiplexed gene expression, automated multidimensional flow cytometry, and integrated single-cell assays in nanowells to quantitatively assess leukocyte function to ultimately identify the molecular signatures defining individual immune responses. We will address immune profiles in three related studies. In Research Project 1, we will develop immunologic signatures of influenza vaccine responsiveness and determine the effect of aging and functional status on these signatures. We will identify gene signatures and biological pathways that can distinguish between strong and weak immune responses to vaccination and that predict effective responses. In Research Project 2, we will investigate resistance to flaviviral infections using West Nile virus, and hepatitis C virus;through analysis of responses in patients from stratified cohorts, we will establish correlations between gene expression, immune cell responses and clinical outcome. In Research Project 3, we will generate mathematical models that detect connectivity and predict dynamic functional responses of the immune system. This approach will link data collected on both populations and individuals using multivariate statistical approaches to integrate cohort-wide data including genome wide association studies and novel single-cell analyses to assess immune responsiveness in relationship to genetic variation. Our functional systems immunology approach will allow us to define baseline human immune signatures following viral infection and vaccination along with deviations from this baseline, with the goal of identifying future targets for intervention and establishing sets of biomarkers that predict responses to vaccination PROJECT 1: Immune Responses Defining Efficiency of Influenza Vaccination Project Leader: Shaw, A PROJECT 1 DESCRIPTION (provided by applicant): The genetic variability of our species dictates that some individuals will develop strong humoral and cellular immune responses to vaccines whereas others do not. With aging, these differences in protective immunity become even more severe, reflected in the increased risk for death and morbidity from infections in older individuals. For example, the impact of seasonal influenza is particularly acute in the geriatric age group, with 90% of the 20 to 40 thousand annual deaths attributed to influenza occurring in individuals over the age of 65. However, the efficacy of the trivalent inactivated influenza vaccine is as low as 30%, and the frail subset of elderly individuals who are at risk for worsened disability, hospitalization, falls and death, represent a particularly vulnerable population. This project builds on our experience recruiting and evaluating influenza vaccine response in young and older individuals, and our access to unique cohorts-such as frail elderly individuals (including a recruited cohort of 860 nursing home elders participating in an NIH-funded trial of pneumonia prevention) and a group of 300 individuals under the age of 30 already subjected to genome wide genotyping. We will utilize the Multidimensional Flow Cytometry and Quantitative Gene Expression Cores, and the analytic methods of Project 3 to develop cellular and gene expression signatures of a successful innate and adaptive immune response to influenza vaccination, and will elucidate the impact of aging and impaired functional status (such as the geriatric syndrome of frailty) on these signatures in cohorts of young, non-frail older, and frail older individuals. Our access to genetic information on 300 genotyped individuals receiving influenza vaccine will also facilitate the integration of cellular and gene expression data with genetic correlates of vaccine response. Identifying the genes and their allelic variations in humans that underlie robust or weak responses, and how their expression patterns are affected by age or frailty is a necessity for a greater understanding of the function of our immune system. Moreover, understanding the genetic architecture of immune responses is likely to identify immune pathways that could be targets of therapies, drugs or other biological treatments to enhance or suppress immune responses as needed. RELEVANCE (See instructions): The goal of this proposal is to identify patterns of gene expression or cell function in the human immune system that are "signatures" of a response to influenza vaccination that is associated with protection from infection. We will also evaluate how these signatures are altered in older or frail individuals, who usually do not generate protective responses to vaccination and are at increased risk for severe influenza infection.